1. Field of the Invention
The present invention relates to a confocal microscope with which a confocal image of a sample to be observed can be obtained.
2. Description of the Related Art
Conventionally, there are microscopes having a confocal optics system (hereinafter referred to as a confocal microscope) that illuminates a sample to be observed with a pointlike light source in a pointlike shape, forms the pointlike image of the light reflected from the illuminated sample, and obtains the density information of the image with a detector via a pinhole.
FIG. 1 is a schematic diagram showing the outline of the configuration of the confocal optics system. As shown in this figure, light irradiated from a pointlike light source 701 gathers at one point of a sample to be observed 704 with an objective lens 703 the aberration of which is properly corrected, and illuminates the sample 704.
The light reflected from the sample 704 is formed as a pointlike image on a pinhole 705 via a half mirror 702, and the light formed as the pointlike image is detected by a light detector 706 via the pinhole 705. By two-dimensionally scanning the sample 704, for example, similar to the raster scanning of a television, a two-dimensional image (hereinafter referred to as a confocal image) is obtained.
Here, reflected light in a position “a” displaced from the light-gathering position of the objective lens 703 is not gathered on the pinhole 705. Therefore, the reflected light does not pass through the pinhole 705 and is not detected by the light detector 706. Accordingly, with the optics system shown in FIG. 1, the image only in the light-gathering position, namely, the focusing position of the objective lens 703 can be obtained.
The focal depth of the above described confocal optics system is shallow. Therefore, there are no problems if an observation surface of the sample 704 is a flat surface. However, if the sample has non-flat observation surfaces where there are projections and depressions of different lengths as shown in FIG. 2, the image of a surface of the sample away from the focusing position cannot be obtained.
FIG. 2 shows an example of the sample having the non-flat observation surfaces. The sample 800 shown in this figure is a sample having non-flat observation surfaces A, B, and C of different heights a, b, and c (b>a>c). Z is the irradiation direction of light. For example, when the observation surface A is focused, observation images of the observation surfaces B and C cannot be obtained.
Accordingly, it is impossible to observe all the observation surfaces A, B, and C by performing an observation operation only once. To observe all the observation surfaces A, B and C, observation images obtained by individually focusing the observation surfaces A, B and C must be synthesized.
However, an observer must determine the focusing position of each of, for example, the observation surfaces A, B and C while viewing an observation image. Therefore, the observer must determine focusing positions for all of observation surfaces whenever he or she observes a sample like that 800 shown in FIG. 2, leading to a heady load on the observer.
As a solution to this problem, Patent Document 1 discloses a scanning-type optical microscope that can obtain an observation image of a sample without determining focusing positions for all of observation surfaces by automatically setting an image capturing range for a sample having the observation surfaces of different heights, and can reduce a load on an observer.
With the scanning-type optical microscope disclosed by Patent Document 1, a confocal image is captured while the microscope moves between first and second stages, and a focused image (herein after referred to as an extended image) in an entire measurement region can be obtained even if a user does not set the image capturing range.
However, according to Patent Document 1, the image capturing range is determined depending on whether or not the number of pixels of a predetermined brightness level or lower is equal to or larger than a preset number of pixels in a confocal image obtained in an arbitrary stage position. Accordingly, for a sample having a plurality of level differences (for example, the sample 800 shown in FIG. 2), the image capturing range is erroneously determined to end at the initial level difference.
As a solution to this problem, Patent Document 2 discloses a scanning-type microscope that can automatically sets an image capturing range without fail even for a sample having observation surfaces of different heights.
The scanning-type microscope disclosed by Patent Document 2 uses the numbers of observation surfaces (the numbers of flat surfaces) of a sample to be observed. Namely, the number of flat surfaces Nup in an upper limit direction of an initial focusing position, and the number of flat surfaces Ndown in a lower limit direction are used. Assuming that the initial focusing position exists between A and C in the sample shown in FIG. 2, the numbers of flat surfaces Nup and Ndown are 2 and 1 respectively.
Then, the number of pixels of brightness equal to or lower than a predetermined level in an obtained confocal image is counted while the stage on which the sample is put is being moved in one direction, and a flat surface is determined depending on whether or not the counted number of pixels is equal to or larger than a predetermined number of pixels. Then, the number of flat surfaces is counted each time a flat surface is detected.
A position where the number of flat surfaces reaches a set value (for example, Nup=2) is stored as a first stage position. Similarly, the number of pixels of brightness equal to or lower than a predetermined level in an observation image is counted while the stage on which the sample is put is being moved in the reverse direction, and a flat surface is determined depending on whether or not the counted number of pixels is equal to or larger than a predetermined number of pixels. Then, the number of flat surfaces is counted, and a position where the number of flat surfaces reaches a preset value (for example, Ndown=1) is stored as a second stage position.
With the above described process, the first and the second stage positions are detected and the confocal image between the first and the second stage positions can be obtained even for a sample having observation surfaces with a plurality of level differences. As a result, an extended image can be obtained.    [Patent Document 1] Japanese Published Unexamined Application No. H06-308393    [Patent Document 2] Japanese Published Unexamined Application No. H08-278450